The major objective of the proposed R21 investigation is to examine the influence of ethanol and ethanol metabolism on major histocompatibility complex (MHC) class l-restricted presentation of antigenic peptides in ethanol-metabolizing liver cell lines. After activation with interferon gamma (IFNy), these peptides are cleaved by major antigen-trimming enzymes, the proteasome and leucine amino peptidase. Our previous investigations have indicated that ethanol suppresses proteasome activity in ethanol-metabolizing HepG2 cells, and that these ethanol-treated cells do not properly transduce the IFNy signal. We hypothesize that in liver cells, ethanol metabolism blocks IFNy-mediated signal transduction. This impairs IFNy-regulated induction of the immunoproteasome and of leucine amino peptidase, compromising their ability to generate peptides for antigen presentation and may alter immune response. We therefore propose the following Specific Aims: SPECIFIC AIM 1: To investigate whether ethanol or its metabolism affects the IFNy-induced cleavage of peptides for MHC class l-restricted antigen presentation in mouse recombinant HepB6 cells and in human recombinant HepG2 cells that express alcohol dehydrogenase and cytochrome P4502E1. SPECIFIC AIM 2: To determine whether ethanol exposure affects IFNy-mediated signal transduction by examining ethanol-elicited alterations in specific components of the JAK-STAT1 signal transduction pathway in mouse recombinant HepB6 cells and in human recombinant HepG2 cells. SPECIFIC AIM 3: To determine whether ethanol or its metabolism affects MHC class l-restricted antigen presentation by liver cells, using a model of the intracellular cleavage of ovalbumin (OVA) and the presentation of OVA peptide SIINFEKL-H-2Kb complex on the surface of mouse recombinant HepB6 cells. The results derived from this study will clarify the link between the effects of ethanol on antigen presentation by liver cells, altered immune response and the possible mechanisms of alcohol-related progression of viral hepatitis. [unreadable] [unreadable]